The Effects And Gene Expression Analysis Of MWCNTs On RAW264.7 Macrophages

Carbon nanotubes (CNTs) are currently considered as one of the important materials in nanotechnology. The potential applications of carbon nanotubes range from biomedicine to mechanical engineering. The number of industrial scale facilities for the relatively low-cost production of MWCNTs continues to grow, and with that the professional and public exposure to MWCNTs is expected to increase significantly in the near future. Thus, it is essential to know the potential hazards of CNTs, particularly MWCNTs, to humans and other biological systems. Thus, it is important to know the potential hazards of carbon nanotubes in vivo and in vitro.In the present study, the aim of this study was to investigate and compare the in vitro responses of RAW264.7 macrophages after exposure to the pristine multi-walled carbon nanotubes(p-MWCNTs) and taurine functionalized multi-walled carbon nanotubes (tau-MWCNTs). We also investigated the regulatory effects of the p-MWCNTs and tau-MWCNTs on RAW264.7 macrophages by using cDNA microarrays.Objective:To study the toxicological effects and mechanisms of the p-MWCNTs and tau-MWCNTs on RAW264.7 macrophages in vitro. Methods:We focused on their effects on cell viability, LDH release, determination of GSH/GSSG ratio, annexin V/PI staining and intracellular calcium concentration, ultrastructural changes in cell morphology, cell cycle, as well as on the release of IL-8 and TNF-a. Results:The present study showed that exposure of RAW264.7 macrophages to p-MWCNTs at high dosage could induce oxidative stress, cellular toxicity and apoptosis, as indicated by the loss of cell viability, increase in LDH release, decline in the cellular GSH/GSSG ratio, increase of IL-8 and TNF-α, arrest in G0/G1, as well as increase of intracellular calcium concentration. Whereas tau-MWCNTs had only very limited impact on oxidative stress, cellular toxicity and apoptosis. Additionally, exposure to p-MWCNTs resulted in ultrastructural and morphological changes in RAW 264.7 macrophages. In contrast, the RAW 264.7 macrophages exposed to the tau-MWCNTs did not exhibit altered morphology.Conclusion:Our results imply that the tau-MWCNTs should be used for biological or medical purposes without affecting the cell viability. It may be used as drug carriers in tumor therapy, fibrillation atrial, cerebrovascular Disease. Maybe someday it can be used generally in clinical application.Technique of cDNA microarray contains molecular biology, material science, microelectronics, physics, chemistry, computer and information science and so on. Gene expression analysis was widely applied in DNA sequencing, the analysis of gene expression, genome, gene diagnosis, the study and development of new medicines. Objective:Using Affymetrix Genechip mouse 430 2.0 array oligonucleotide arrays, we had studied the possible mechanism of p-MWCNTs and tau-MWCNTs on RAW264.7 macrophages. Methods:Changes of gene expression in RAW264.7 macrophages, after treatment with p-MWCNTs and tau-MWCNTs, were examined by the cDNA microarray in order to study the molecular mechanism. Real time RT-PCR was conducted to verify the differential expression of some genes. Results:After the RAW 264.7 macrophages were exposed to the p-MWCNTs for 24h, compared with the control, there were 5738 genes showed differential expression, of which 1905 genes were up-regulated and 3833 genes were down-regulated. On the other hand, after the RAW 264.7 macrophages were exposed to the tau-MWCNTs for 24h, compared with the control, there were 2567 genes showed differential expression, of which 959 genes were up-regulated and 1608 genes were down-regulated. According to the Gene Ontology, these genes were involved in cell cycle, apoptosis, stress and inflammation, cytoskeleton, intracellular transport, nitric oxide metabolic process, DNA repair, DNA-binding transcription, energy metabolism, Ion channels and transport proteins, cell signaling, cell receptor and so on. Conclusion:The p-MWCNTs inhibit the proliferation of the RAW 264.7 macrophages by regulating the genes expression, including cell cycle, apoptosis, stress and inflammation and so on, of which the mechanisms are complicted. There are reduced differential expression genes on the RAW 264.7 macrophages after treatment with the tau-MWCNTs. We analyze some genes including cell cycle, apoptosis, stress and inflammation, which are confirmed by the real time RT-PCR, and the results are corresponded to that of cDNA microarray.